PURIFICATION AND SOME PROPERTIES OF MALTO SE PHOSPHORYLASE FROM ENTEROCOCCUS-HIRAE IFO-3181

Maltose phosphorylase was isolated to homogeneity from a cell-free ext ract of Enterococcus hirae IFO 3181 by chromatographies with phenyl-Se pharose CL-4 B, QAE-Sephadex A-50, hydroxylapatite, and Sephadex G-200 . The enzyme was purified about 40-fold with a yield of 32%, and its s pecific activity was 29.4 units/mg protein. The molecular weight was e stimated to be 220,000 and 90,000 by HPLC gel filtration on TSKgel G 3 000 SWXL and SDS-polyacrylamide gel electrophoresis, respectively. The enzyme showed optimum activity around pH 6.5-7.5 and its optimum temp erature was about 45 degrees C. The enzyme was stable over the range o f pH 5.5-8.0 and retained the activity up to about 55 degrees C. Malto se and nigerose were effective substrates for the enzyme reaction, but it could not act on the other disaccharides tested. The Km for maltos e, nigerose, phosphate, and arsenate were 1.90, 1.93, 3.37, and 8.33 m M, respectively. The enzyme activity was almost completely inhibited b y AgNO3 and HgCl2, and also strongly inhibited by CuSO4, ZnSO4, and p- chloromercuribenzoic acid.